Organophosphate (OP) anticholinesterases, e.g., nerve agents or insecticides, are potent acetylcholinesterase (AChE) inhibitors. High dose poisoning causes excitotoxicity that leads to seizures and subsequently brain damage. The current therapy in the US includes an oxime reactivator, 2-PAM, which has little, if any, ability t cross the blood-brain barrier (BBB) and consequently cannot stop seizures and prevent brain damage. Our laboratories have developed a library of phenoxyalkyl pyridinium oximes designed to have greater lipophilicity than 2-PAM. Through a paradigm using high sub-lethal dosages of highly relevant surrogates for sarin or VX in rats and administering the novel oximes at 1 hr after OP compound treatment when seizure behavior was on- going, we have demonstrated the ability of a number of these oximes to penetrate the BBB, reactivate AChE, and to attenuate seizure behavior. The objective of this application is to further characterize a limited number of these novel oximes (initially 6, then down-selected to 3) to ultimately select a single lead oxime for an IND application. Additionally we will expand the OP chemistries tested to include an insecticidal chemistry, paraoxon (the active metabolite of parathion; tested first to avoid any confounders that could occur because of the need for bioactivation) and parathion. Endpoints of characterization include brain and peripheral AChE reactivation, seizure attenuation, neurodegeneration (glial fibrillary acidic protein accumulation), oxidative stress (F2-isoprostanes), neuropathology (apoptosis, neuron survival), and behavior. Concurrently molecular dynamics computations will determine if one or two additional oxime structures are predicted to have greater efficacy than our current selections; if so, it/they will be synthesized and substituted for our current selections. The specific aims of the proposed project are: 1. In vivo efficacy characterization aim: To provide expanded measures of neuroprotection on our currently identified BBB-penetrating oximes using the sarin and VX surrogates and also a common insecticidal OP chemistry (i.e., diethyl phosphate) using parathion and its active metabolite paraoxon. Efficacy endpoints for the 4 OP's include: 1a, efficacy screening endpoints with 6 oximes: reactivating rat brain AChE, and the attenuation of seizure behavior and neurodegeneration; and 1b, detailed efficacy characterization endpoints with 3 down-selected oximes: oxidative stress in the brain, neuropathology and behavioral deficits. 2. Pharmacokinetic aim: To determine for the 3 down-selected oximes hepatic in vitro metabolism, plasma protein binding and levels of oxime in the blood and cerebrospinal fluid (obtained through microdialysis). 3. Computational chemistry aim: To model the docking of oxime with AChE and BBB penetration to suggest improved oxime structures. 4. Chemical synthesis aim: To produce sufficient quantities of the test OP's and the test oximes for the experiments proposed, as well as the synthesis of a few new oximes if suggested by the computational chemistry aim. The ultimate outcome will be the identification of the most efficacious novel oxime for further development as a potential substitute for current therapy.